Hello,
I'm doing PCR and gel electrophoresis on tetranucleotide microsatellite repeat of a species of diploid salamander. The gel results yield three bands in two sample (attached; red arrows), and I'm confused about how should I interpret this. Other samples (not shown) also display three bands. I only expect from one to two bands in diploid organism. There are a lot of non-specifics, but the microsatellite alleles are highly pronounced.
Thank you
Hi!
There are reports available on tri-allelics. Go through the following and attached. You may get some information. Good Luck!!
tri-allelic inheritance
(https://genetics.thetech.org/ask-a-geneticist/triallelic-diseases-and-inheritance)
Observation of tri-allelic patterns in autosomal STRs during routine casework
(https://www.fsigeneticssup.com/article/S1875-1768(09)00003-1/pdf)
Identification of a functionally significant tri-allelic genotype in the Tyrosinase gene (TYR) causing
hypomorphic oculocutaneous albinism (OCA1B)
(https://www.nature.com/articles/s41598-017-04401-5.pdf)
Assessment of intra and interregional genetic variation in the Eastern Red-backed Salamander, Plethodon cinereus, via analysis of novel microsatellite markers
(https://journals.plos.org/plosone/article/file?id=10.1371/journal.pone.0186866&type=printable)
Tri-allelic pattern at the TPOX locus: human identification in Brazil
( https://www.oatext.com/pdf/FSC-3-127.pdf )
Tri-allelic patterns at the D7S820 locus detected in two generations of a Chinese family
(https://link.springer.com/content/pdf/10.1007%2Fs00414-015-1166-6.pdf)
Hello
Well, there could be several things. They may be nonspecific amplifications, if so, you would have to perform tests to improve the PCR (eg raise the anneling temperature), with microsatellites it is relatively normal that you have to standardize per sampling site. Another option is that you effectively have more than two alleles.
One way you could make sure is to perform the capillary electrophoresis in a sequencer, and having the results and according to the shape of the alleles you can know if they are indeed alleles or stutter.
- Badges
- Science method